Abstract A prospective, randomized, crossover experimental pilot study was performed to compare the anesthetic quality and recovery times of intramuscular (IM) ketamine/medetomidine and S-ketamine/medetomidine in Testudo spp. Four adult male Hermann's tortoises (Testudo hermanni) and one adult male spur-thighed tortoise (Testudo graeca) were used in this study. Each animal underwent two different anesthetic protocols twice, with a washout period of two weeks in between each anesthetic event. This resulted in 20 anesthetic procedures. Protocols consisted of 0.1 mg/kg medetomidine IM with either 15 mg/kg ketamine (KM) or 15 mg/kg S-ketamine (SKM). After 60 min, medetomidine was reversed using 0.5mg/kg atipamezole IM. Muscle relaxation, ease of handling, sensitivity to a painful stimulus, palpebral and corneal reflexes, heart rate (HR), and respiratory rate (RR) were measured every 15 min during anesthesia. Time of initial effect, maximal anesthetic effect, first movements, complete wakefulness, first feedin...